Monolithic column electroelution apparatus and method thereof

An electric elution and monolithic column technology, applied in the field of analytical chemistry, can solve the problems of low recovery rate, long elution time, cumbersome operation, etc., and achieve the effects of simple production, shortened elution time, and simplified operation steps

Inactive Publication Date: 2012-12-12
SHANGHAI JIAO TONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The purpose of the present invention is to provide a protein or nucleic acid monolithic column electroelution device and its method for the above existing problems such as long elution time, low recovery rate and cumbersome operation, so as to obtain the maximum recovery rate in the shortest time and also Different samples can be eluted

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  • Monolithic column electroelution apparatus and method thereof
  • Monolithic column electroelution apparatus and method thereof
  • Monolithic column electroelution apparatus and method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Example 1: Electroelution of Target Proteins from Agarose Gel Electrophoresis (AGE)

[0034] Step 1: Perform protein (such as bovine serum albumin) AGE

[0035] Prepare agarose gel (0.8 % agarose, w / v) with 9 mM barbiturate buffer (pH 8.6). The sample was dissolved in 5 ml of 9 mM barbiturate buffer (pH 8.6) with standard bovine serum albumin 5 mg, and it was completely dissolved by stirring. Then add 160 microliters of sample solution to load the sample for agarose electrophoresis, the electrophoresis voltage is 100 V, and the experimental temperature is 25 o C, the electrophoresis time is 35 min. After electrophoresis, take out the gel, cut off both sides of the gel for quick staining, and put it back on the original gel plate to locate the unstained protein band after completion.

[0036] Step 2: Cut the gel plate containing the bands of the protein of interest (such as albumin) and place it in the slot

[0037] Cut according to the position of the target prote...

Embodiment 2

[0047] Example 2: Electroelution of Target Proteins from Polyacrylamide Gel Electrophoresis (PAGE)

[0048] Step 1: Perform protein (such as serum protein) PAGE

[0049] PAGE electrophoresis conditions: polyacrylamide separating gel (T: 10%, C: 2.7%), polyacrylamide stacking gel (T: 4%, C: 2.7%), 1500 mM Tris-Hcl separating gel buffer (pH 8.9 ), 500 mM Tris-Hcl stacking gel buffer (pH 6.8), 25 mM Tris-Gly electrode buffer (pH 8.3), serum samples were dissolved in 50 mM Tris-Hcl stacking gel buffer (pH 6.8, containing 1% bromine phenol blue), 10-fold dilution, 10 microliters of sample per well, electrophoresis at 60 V until the bromophenol blue enters the separation gel, and then electrophoresis at 160 V until the bromophenol blue reaches the lower end of the gel, and the electrophoresis time is 90 min. After electrophoresis, take out the gel, cut off both sides of the gel for quick staining, and put it back on the original gel plate to locate the unstained protein band afte...

Embodiment 3

[0060] Example 3: Electroelution of Target Nucleic Acids from Agarose Gel Electrophoresis (AGE)

[0061] Step 1: Perform nucleic acid AGE

[0062] Carry out DNA agarose electrophoresis according to conventional conditions, the gel concentration is 0.5%, and the gel contains EtBr 0.5 μ g / ml, the electrode buffer is TAE (40 mM, pH 8.0), and the voltage is 4V / cm. After electrophoresis, the gel was moved to the UV lamp (360 nm), and the target band was quickly cross-cut with an autoclaved sterile scalpel blade, and then the UV lamp was turned off for subsequent operations under visible light.

[0063] Step 2: Put the cut gel plate containing the nucleic acid zone into the slot

[0064] According to the target nucleic acid band drawn in step 1, remove and put the complete gel plate into the slot, the width and thickness of the gel plate match the slot, and cling to the small cylinder on the other side.

[0065] Step 3: Put the dialysis tubing on the socket and add the eluent ...

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Abstract

The invention relates to a monolithic column electroelution apparatus and a method thereof. The monolithic column electroelution apparatus comprises an electrophoresis tank with electrodes, a gel plate containing a target protein zone or a nucleic acid zone, slots, a dialysis tube, and a frame, wherein the gel plate is placed in the slot, the dialysis tube is sleeved outside the slot, both ends of the dialysis tube are folded upward, the slot is placed inside the frame, and long axis of the slot is parallel to the electrodes on the outer sides of the frame. According to the present invention, the difficult problem of sample recovery is overcome by assembling simple materials in the experiment, such that characteristics of high sample recovery rate and short elution time are provided, elution of a plurality of samples can be concurrently performed, operation is simple, and the method of the present invention can be adopted as an ideal elution and pre-enrichment method.

Description

technical field [0001] The invention relates to the technical field of analytical chemistry, in particular to an integral column electroelution device and method for recovering protein or nucleic acid from gel. Background technique [0002] The purification and recovery of samples is a routine operation in biochemical laboratories and a key step in the process of protein or nucleic acid analysis. In the process of protein or nucleic acid analysis, it is often necessary to separate and extract specific proteins or nucleic acids from the gel for subsequent operations (such as SDS-PAGE, LC-MS and MS analysis and identification, etc.). Therefore, the recovery of protein or nucleic acid bands after gel electrophoresis directly affects the process, results, and even success of the entire protein or nucleic acid separation and analysis process. [0003] At present, there are mainly four elution methods for the recovery of protein or nucleic acid bands in electrophoresis. The firs...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K1/24C07H1/06
Inventor 曹成喜李国庆邵菁樊柳荫郭陈刚刘小平
Owner SHANGHAI JIAO TONG UNIV
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